(A) Field of the Invention
This invention relates to the preparation of aqueous dispersions of water-swellable polymer particles and to the particles obtained from such a process.
(B) Description of the Prior Art
Aqueous dispersions of water-soluble polymer particles are difficult to manufacture. It is particularly difficult to control particle size and shape and to make such particles reproducibly. Aqueous dispersions of water-soluble polymer particles that have been available prior to this invention are, in general, not satisfactory. Previously, such particles are generally prepared by adding a crosslinking agent to an aqueous polymer solution under agitation. Agitation serves to break down the crosslinking polymer to give the desired particle size. To break down the crosslinked and crosslinking polymers, sufficient mechanical stress to cause shearing and disruption of particle integrity is required. This level of mechanical stress can only be achieved by applying vigorous agitation. This concurrent agitation-crosslinking method gives an undesirably broad distribution of particle sizes and shapes, and produces undesirable particle fragments. Additionally, there is substantial variability in the resulting particle populations between production runs.
A particularly undesirable aspect of the heterogeneous particle population produced by this method is that it is difficult or impossible to xe2x80x9cclean upxe2x80x9d or fractionate the heterogeneous population to yield a homogeneous population of particles having the same size and shape. A homogeneous population of particles is particularly desirable where the particles are to be delivered by injection through a narrow guage syringe needle.
Other processes for preparing aqueous dispersions of crosslinked water-soluble polymer particles involve methods that are of a proprietary nature. However, such particles are either not available in the quantities to meet commercial demand, their physical parameters are not suitable for a particular application, or their properties cannot be varied systematically. Moreover, a full range of polymer compositions and sizes has not been available.
Water-soluble particles have various utilities. There are three basic types of water-based polymers:
(a) solution polymers. These polymers comprise dispersions of individual polymer molecules. The viscosity of the solution depends upon the molecular weight and concentration of the polymer; the higher the molecular weight and concentration, the higher is the viscosity of the solution. The viscosity of the solution limits the use of these solution polymers to low molecular weights and low concentrations. This is especially true where the use involves delivery of particles via injection through a narrow gauge syringe needle.
(b) latexes. These polymers comprise colloidal dispersions of polymer particles, each of which comprises hundreds or thousands of polymer molecules. The viscosity of the latex depends upon the interactions between the colloidal particles and is independent of the molecular weight of the polymer. Thus, latexes often combine low viscosities with high polymer concentrations. Moreover, the mechanism and kinetics of emulsion polymerization favor the preparation of high-molecular-weight polymers with rapid rates of polymerization. Aqueous dispersions of crosslinked water-soluble polymers can be prepared by inverse emulsion or inverse suspension polymerization of monomer mixtures containing a crosslinking monomer, e.g., a mixture of acrylamide and methylene-bis-acrylamide. However, this method is limited to polymers that can be prepared by radical chain polymerization, which excludes natural water-soluble polymers. There remains a need to prepare particles of water-soluble polymers that are derived from natural sources.
(c) Water-reducible or water-dispersible polymers. These polymers have a degree of dispersion intermediate between that of solution polymers and that of latexes. The viscosity of these intermediate samples depends upon the relative degrees of solution polymer and latex polymer character. As with the latexes, this method is limited to polymers that can be prepared by radical chain polymerization.
Polysaccharides are naturally occurring biopolymers that exist in a highly viscous liquid state in animal tissues, where they readily react with proteins to from glycosaminoglycans or proteoglycans.
Biocompatible and non-biodegradable particles that are non-cytotoxic, non-carcinogenic, non-inflammatory, non-pyrogenic, and non-immunogenic are needed to provide a solution to the long felt and unfulfilled need for an improved composition useful for implants; soft tissue augmentation to treat congenital abnormalities, acquired defects, and cosmetic defects; and tissue scaffolding to promote cell growth.
Homogeneous populations of such particles would be particularly useful for soft tissue augumentation by surgical implantation or, preferrably, delivery to the desired site by conventional injection through a narrow gauge syringe needle.
It would be particularly desirable to use such particles to treat urinary incontinence and vesicoureteral reflux, for correction of wrinkles and other skin defects, or to serve as a general augmention or replacement composition or as a scaffold material in soft or hard tissues such as breast, lip, penis, bone, cartilage, and tendon.
U.S. Pat. No. 4,124,705 to Rothman, et al. (hereinafter xe2x80x9cRothmanxe2x80x9d) discloses hydrophilic, water-insoluble particles having a particle size between 0.1 and 300 micrometers which are composed of a polysaccharide or polysaccharide derivative such as starch, glycogen, or dextrins. The particles are crosslinked into a three-dimensional network by xcex1(1xe2x86x924)glucosidic linkages. This network is degraded in the body through hydrolyzation of the xcex1(1xe2x86x924)glucosidic linkages by xcex1-amylase to form water-soluble fragments.
The Rothman particles are produced by bead polymerization in which a solution of the polysaccharide is dispersed to droplet form in an inert liquid, such as octanol, which may contain an emulsion stabilizer, such as Gafac(copyright) PE 510. A crosslinking agent, for example, a di- or multi-epoxides or a dicarboxylic acid, is then added to the reaction mixture. The octanol used in the Rothman process is a polar solvent which is miscible with the aqueous solution of the polysaccharide.
The Rothman particles are administered intravascularly in aqueous solutions, such as glucose, sorbitol, or saccharose, to block the finer blood vessels leading to a particular part of the body. This prevents the tissue in that part of the body, for example, a tumor, from receiving necessary oxygen and nutrients and inhibits growth of the tissue. The particles may also be administered with a diagnostic or therapeutic agent, allowing the agent to be trapped within or without of the effected tissue for brief periods of time.
European patent application 256,293 to Mitsubishi discloses water-insoluble, crosslinked, polyvinyl alcohol particles in the shape of spheres having an average particle size from 20 to 1,000 micrometers. The particles are produced by dispersing a solution of polyvinyl alcohol and a salt in an organic solvent, such as a hydrocarbon, to form a gel which is then reacted with a crosslinking agent, such as a dialdehyde, diepoxide, glycidyl ether, and epihalohydrin. Suitable salts are sodium chloride, sodium sulfate or any other salt capable of coagulating and precipitating polyvinyl alcohol. A dispersion stabilizer, such as a cellulose or sorbitan derivative, may be added to the reaction mixture. Mitsubishi further discloses that the crosslinked polyvinyl alcohol particles are suitable for packing materials in chromatography.
European patent application 555,980 to Nisshinbo discloses crosslinked, spherical particles of water-soluble polymers, having a particle size from 0.1 to 30 micrometers. Specific polymers disclosed are sodium alginate, dextran, dextran sulfate sodium, carragheenan, agarose, agar, gelatin, pectin, water-soluble cellulose derivatives, such as carboxymethylcellulose sodium. The polymer particles also contain an oligosaccharide or polyhydric alcohol, such as sucrose, which is necessary to provide the particles with a spherical shape. Specific oligosaccharides disclosed are mannose, sucrose, cellobiose and raffinose; specific polyhydric alcohols are polyethylene glycol or erythritol. The Nisshinbo particles are useful as additives and binders because of their water-retaining and lubricating properties.
The polymeric particles of Nisshinbo are made by preparing an aqueous solution of the water-soluble polymer and an oligosaccharide or polyhydric alcohol. This solution is then spray-dried to form spherical particles. Nisshinbo discloses that it is not possible to obtain spherical particles through spray drying techniques without the addition of an oligosaccharide or polyhydric alcohol. The spray dried particles are then crosslinked. Specific covalent crosslinking agents disclosed are divinyl compounds or bisepoxide, while specifically disclosed ionic crosslinking agents are calcium chloride and other divalent metal salts.
U.S. Pat. No. 4,716,154 to Malson, et al. (hereinafter xe2x80x9cMalsonxe2x80x9d) discloses a transparent, homogeneous, crosslinked hyaluronic acid gel which is a clear optical mass useful for replacement of vitreous humor in individuals with retinal detachment. The gel may contain other polysaccharides in additional to hyaluronic acid. The gel is administered by injection through a 0.9 millimeter needle tip.
The process by which the gel of Malson is made involves dissolving hyaluronic acid and a crosslinking agent in an alkaline medium, preferably at an elevated temperature of about 50xc2x0 C. The resulting gel must be washed to remove unreacted crosslinking agent. The prefered crosslinkers disclosed are di- or polyfunctional epoxides.
U.S. Pat. No. 5,603,956 to Mateescu, et al. (hereinafter xe2x80x9cMateescuxe2x80x9d) discloses crosslinked polymer particles of amylose having a size of about 0.5 to about 5.0 micrometers which form agglomerates of approximately 25-700 micrometers. The polymers are crosslinked solely through xcex1(1xe2x86x924) linkages.
The Mateescu particles are formed by direct compression of an admixture of a drug with the crosslinked amylose polymer and an amount of xcex1-amylase enzyme. The crosslinked amylose polymer is formed by swelling amylose in an alkaline medium in a planetary mixer, with homogenization, followed by addition of the crosslinker with moderate heating of between 40-70xc2x0 C.
The polymers are useful for the slow release of drugs. The xcex1-amylase present in the particles breaks down the xcex1(1xe2x86x924) linkages, releasing the drug and degrading the polymer.
U.S. Pat. No. 5,371,208 to Kozulic discloses electrophoresis gels having a very low polymer concentration and improved optical properties. The gels are formed by reacting a hydroxyl group containing polymer with a crosslinker that is capable of forming ether linkages, such as bis-epoxides, in an aqueous medium at a basic pH. Because the process is performed in water, hydrolysis of the reactive groups present in the crosslinker is an unavoidable side reaction.
U.S. Pat. No. 5,041,292 to Feijen discloses drug delivery system containing a drug and a biodegradable hydrogel matrix, formed by linking a polysaccharide and a protein with a crosslinking agent. The Feijen process involves dissolving the polysaccharide, protein, and crosslinking agent in an aqueous medum. The crosslinked particles are then loaded with the desired drug. The hydrogel may be shaped into many forms, including microspheres which may be from less than 100 nanometers to over 7 micrometers in diameter. Feijen discloses that the size may be varied in order to place the gels in the capillary bed of the lungs, the liver and spleen through phagocytosis of small particles, and in the extracellular tissue.
The particles formed by Feijen are maintained by mechanical agitation prior to crosslinking, and once formed are stabilized by heat, which is only possible in the presence of the additional protein component. Thus, the size and shape of the particles are controlled solely by the mechanical force of the system.
Thus, there is a need for aqueous dispersions of water-swellable polymer particles of a relatively narrow particle size distribution with defined physical characteristics, for a process for preparing such dispersions, and for recovery of particles in any desired quantity at reasonable cost.
Stated generally, the invention comprises in one embodiment a process for producing a crosslinked water-swellable polymer particle preparation and the particles produced by the process. In the first step of the process of the invention, an aqueous polymer solution containing at least one water-soluble polymer, having at least one functional group or charge, is combined with an aqueous medium. The aqueous polymer solution is then mixed under moderate agitation with an oil phase containing an inert hydrophobic liquid and at least one emulsifier to form an emulsion of droplets of the water-soluble polymer. At least one crosslinking agent capable of crosslinking the functional groups and/or charges in the water-soluble polymer is then added to the emulsion to form crosslinked water-swellable polymer particles. This emulsion of crosslinked water-swellable polymer particles may optionally be inverted in an excess of water to provide an aqueous dispersion of crosslinked water-swellable polymer particles. Optionally, the crosslinked water-swellable polymer particles can be recovered from the aqueous dispersion by conventional methods.
In one embodiment, the dislosedprocess comprises forming an aqueous solution of a water-soluble polymer having at least one of the following functional groups: hydroxyl groups, thiol groups, carboxyl groups, sulfonic acid groups, sulfate groups, phosphate groups, amino groups, aldehyde groups, and sulfonyl halide groups. This aqueous solution is then added to an oil phase containing a hydrocarbon and an emulsifier having a low HLB value, preferably less than 8.
In another embodiment, the invention generally comprises a crosslinked water-swellable polymer particle preparation. The preparation contains crosslinked water-swellable polymer particles that are substantially homogeneous in size. The particles have a size between about 10 and about 250 micrometers in diameter. The particles are preferably less than 212 micrometers in diameter. At least about 80% of the particles in the preparation are spherical. More specifically, the particle preparation comprises particles composed of one or more water-soluble polymers having at least one of the following functional groups: hydroxyl groups, thiol groups, carboxyl groups, sulfonic acid groups, sulfate groups, phosphate groups, amino groups, aldehyde groups, and sulfonyl halide groups, and at least 95% of the particles are spherical.
In yet another embodiment, the invention generally comprises aqueous dispersions containing crosslinked water-swellable polymer particles that are substantially homogeneous in size. The particles in the dispersion have a particle size between about 10 and about 250 micrometers in diameter, preferably less than 212 micrometers. More specifically, the aqueous dispersions are suitable for medicinal purposes.
Stated generally, another embodiment of the invention comprises the use of crosslinked water-swellable polymer particles and aqueous dispersions containing them for medicinal purposes. More specifically, these methods include administering an aqueous dispersion containing the crosslinked water-swellable polymer particles to an individual as implants, as scaffold material for cell growth, or for soft tissue augmentation. Even more specifically, the invention comprises a method of soft tissue augmentation useful for the treatment of urinary incontinence, vesicoureteral reflux, glottic insufficiency, gastroesophageal reflux, or skin defects. The method also specifically comprises a method of providing scaffolding material for wound healing and for tissue replacement in tissues in the breast, lip, penis, bone, cartilage, and tendon.
Therefore, it is an object of the present invention to provide a simple and cost effective method of preparing crosslinked water-swellable polymer particles.
It is another object of the invention to provide a process for preparing crosslinked water-swellable polymer particles which provides an improved method for controlling the size and shape of the particles formed.
It is yet another object of the invention to provide a process for the production of crosslinked water-swellable polymer particles that provides a high degree of crosslinking.
It is a further object of the invention to provide a process for the production of aqueous solutions of crosslinked water-swellable particles.
It is an object of the invention to provide crosslinked water-swellable polymer particles that are substantially homogeneous in size.
It is another object of the invention to provide crosslinked water-swellable polymer particles having a size between about 10 and about 250 micrometers.
It is yet another object of the invention to provide crosslinked water-swellable polymer particles having a substantially uniform, spherical shape.
It is a further object of the invention to provide crosslinked water-swellable polymer particles containing one or more water-soluble particles having at least one or more functional groups or charges.
It is an object of the invention to provide crosslinked water-swellable polymer particles which are rigid and elastic.
It is another object of the invention to provide aqueous dispersions of crosslinked water-swellable polymer particles which can be injected through a narrow gauge hypodermic syringe needle.
It is yet another object of the invention to provide a method of soft tissue augmentation by administering an aqueous dispersion of crosslinked water-swellable polymer particles.
It is a further object of the invention to provide scaffolding material to promote cell growth by administering an aqueous dispersion of crosslinked water-swellable polymer particles.
The present invention relates to a process for preparing crosslinked water-swellable, hydrophilic polymer particles from water-soluble polymers, the polymer particles formed by the process, and aqueous dispersions and pharmaceutically acceptable dispersions of the polymer particles. The invention also relates to therapeutic methods of using of the polymer particles.
The present invention provides biocompatible and non-biodegradable particles that are substantially non-cytotoxic, non-carcinogenic, non-inflammatory, non-pyrogenic, and non-immunogenic, and which lack other unwanted humoral or cellular responses. The particles also possess sufficient long-term stability of size, shape, rigidity, and compositon to have utility as implant materials. A further characteristic of the particles of the invention which makes them useful for implant purposes is that they are relatively inert and do not rapidly degrade in vivo. A particularly advantageous feature of the particles of the invention is that they are easily injectable. Preferably, the crosslinked water-swellable polymer particles prepared according to the invention are substantially homogeneous in size. They are generally from about 10 microns in diameter to about 250 microns in diameter, and at least 80% of the particles are spherical. Preferably, the particles are greater than about 10 micometers in diameter and less than about 212 micrometers in diameter, and at least 90% of the particles are spherical. In this application, the term substantially homogeneous means that at least 80% of the particles are within one standard deviation of the mean or average size of the particles. Such particles provide a solution to the long felt and unfulfilled need for an improved composition useful for medical treatments, such as implantation; soft tissue augmentation to treat congenital abnormalities, acquired defects, or cosmetic defects; and tissue scaffolding to promote cell growth and wound healing.
Examples of congential defects treatable with the particles of the invention include, without limitation, hemifacial microsomia, malar and zygomatic hypoplasia, unilateral mammary hypoplasia, pectus excavatum, pectoralis agenesis and velopharyngeal incompetence secondary to cleft palate repair or submucous cleft palate. Examples of acquired defects include, without limitation, post surgical, post traumatic and post infectious defects, such as depressed scars, subcutaneous atrophy, acne pitting, linear scleroderma with subcutaneous atrophy, saddle-nose deformity, Romberg""s disease, and unilateral vocal cord paralysis. Cosmetic defects include, without limitation, glabellar frown lines, nasolabial creases, circumoral geographical wrinkles, sunken cheeks and mammary hypoplasia.
The homogeneous populations of particles produced by the process of the invention are useful for soft tissue augumentation involving delivery to the desired implantation site by conventional injection through a narrow gauge syringe needle (such as a 20, 21 or 22 guage needle), although they are also useful for surgical implantation and other delivery methods, such as endoscopy.
The particles of the invention are useful for the treatment of urinary incontinence, vesicoureteral reflux, glottic insufficiency, and gastroesophageal reflux, and for correction of wrinkles and other skin defects, or to serve as a general augumention or replacement composition or as a scaffold material in soft or hard tissues such as breast, lip, penis, bone, cartilage, and tendon. When the particles of the invention are administered to a patient to serve as scaffold material, the particles facilitate the migration and infiltration of fibroblasts and related cells. The large free spaces within the microsphere of the present invention, particularly polysaccharide microspheres, allows cells to infiltrate into and through the bead. Additionally, the large surface area of the microspheres promotes attachment and growth of infiltrating cells. Still further, the scaffolding material is gradually degraded and absorbed by the host. Optionally, when the microspheres of the present invention are used as scaffolding material, bioactive agents can be crosslinked, coupled or otherwise attached, using methods well known in the art, to provide localized cellular stimuli. Examples of such bioactive agents include growth factors and cytokines, such as fibroblastic growth factor, endothelial growth factor, interlukins, platelet derived growth factor, tissue necrosis factor, hormones, cell adhesion peptides, etc. Accordingly, when the microspheres of the present invention are used as scaffolding material, colonization of the site by cells is facilitated.
A particularly advantageous feature of the invention is that process parameters such as the composition of the oil phase employed, the emulsifying agents selected, temperature, pH, crosslinking time and washing, can be adjusted and precisely controlled in small scale synthesis, and then readily and reproducibly duplicated when scaled-up to full production runs to achieve superior microparticles having beneficial characteristics, such as substantially uniform size and shape, excellent physical and chemical stability, and elastic properties that permit easy extrusion through small gauge syringe needles.
The process of preparing crosslinked water-soluble polymer particles comprises dissolving at least one water-soluble polymer having at least one functional group or charge in water or aqueous buffer to provide the desired concentration of the polymer in solution. The aqueous polymer-containing solution is then added to an oil phase which includes at least one water-in-oil emulsifying agent in an amount suitable to provide the desired concentration of the water-soluble polymer. The mixture is agitated moderately to form an emulsion containing droplets of the water soluble polymer. The polymer droplets are crosslinked in-situ by at least one crosslinking agent, resulting in the formation of crosslinked water-swellable polymer particles. Following crosslinking, the polymer-in-oil emulsion is then optionally inverted in an excess of water to provide an aqueous dispersion of the crosslinked polymer particles. The particles may then be recovered from the aqueous dispersion. Preferably, the particles of the invention are formed from a single water-soluble polymer, although particles containing multiple water-soluble polymers are within the scope of the present invention.
Any water-soluble polymer, natural or synthetic, can be used in the practice of the invention provided that (1) the viscosity of an aqueous solution of the polymer is low enough that it can be broken down into droplets during the emulsification step, and (2) it contains at least one functional group or charge that can serve as sites for reaction with a crosslinking agent. Generally, solutions with viscosities as high as 104-105 cps can be broken down into droplets by the usual methods of emulsification.
Water-soluble polymers useful in the invention will contain one or more types of functional groups or charges which can react with a crosslinking agent to form covalent and/or ionic bonds. Examples of functional groups, without limitation, are hydroxyl groups, thiol groups, carboxyl groups, sulfonic acid groups (SO3H), sulfate groups (OSO3H), phosphate groups (OPO3H), amino groups, aldehyde groups, and sulfonyl halide groups (SO2X, where X is Cl or Br).
Examples of water-soluble polymers useful in the process of the invention include, but are not limited to, proteins, polysaccharides, peptidoglycans (heteroglycan chains comprising alternating units of N-acetylglusocsamine (GlcNAc) and N-acetylmuramic acid (MurNAc) linked to various peptides), glycoproteins (proteins to which carbohydrate chains are attached), proteoglycans (proteins to which glycosaminoglycan chains are linked), teichoic acids, lipopolysaccharides, synthetic hydrophilic polymers, and mixtures thereof.
Other hydrophilic polymers useful in the invention include, but are not limited to, natural polysaccharides, such as hyaluronic acid, sodium alginate, chondroitin sulfate, celluloses, chitin, chitosan, agarose, xanthans, dermatan sulfate, keratin sulfate, emulsan, gellan, curdlan, amylose, carrageenans, amylopectin, dextrans, glycogen, starch, heparin sulfate, and limit dextrins and fragments thereof; synthetic hydrophilic polymers, such as poly(ethylene oxide), poly(vinyl alcohol), and poly(N-vinyl pyrrolidone); and proteins, such as bovine serum albumin and human gamma globulin. Polysaccharides are the prefered water-soluble polymers for use in the invention. Particularly, hyaluronic acid and sodium alginate are polysaccharides that are biocompatible and biodegradable in human tissue. They are non-cytotoxic, non-carcinogenic, non-inflammatory, non-pyrogenic, and non-immunogenic, and, therefore, in the form of gel microspheres, they are good candidates for delivery of drugs in accordance with the present invention.
Many crosslinking agents are known in the art. Some form covalent bonds with various functional groups, while others form ionic bonds. The present invention contemplates the formation of polymer particles which are crosslinked through covalent bonds, ionic bonds, or both.
Selection of the particular crosslinking agent for use in the process of the invention will depend upon the particular functional groups present in the water-soluble polymer to be crosslinked. For example, it is known that glutaraldehyde crosslinks amino groups and that XAMA-7 crosslinks carboxyl groups. Divinyl sulfone and epichlorhydrin crosslink hydroxyl and amino groups, and carbodiimides crosslink amino and carboxyl groups. Many epoxides, for example, EGDE and BDDE, crosslink hydroxy groups. The selection of an appropriate crosslinking agent can readily be accomplished by those of skilled in the art.
Examples of crosslinking agents which will be found satisfactory in the present invention include, without limitation, pentaerythritol-tris-[beta (N-aziridinyl)-propionate], divinyl sulfone, XAMA-7, epichlorhydrin, glutaraldehyde, p-toluene sulfonic acid, carbodiimides, epoxides, especially di- and polyepoxides, and ammonium persulfate. These crosslinking agents form covalent bonds with the water-soluble polymers in the process of the invention.
Ions of various alkali metals, alkaline earth metals, and transition metals can also be used to crosslink the water-soluble polymers employed in the present invention. Examples of these metals include, but are not limited to, calcium, magnesium, sodium, potassium, chromium, iron, copper, and zinc. For example, the calcium ion is known to crosslink water-soluble polymers that contain carboxyl groups. These crosslinking agents form ionic bonds with the water-soluble polymers in the process of the invention.
Ionic bonds may be broken down by a change in external conditions, e.g., by chelating agents. On the other hand, covalent bonds are stable in the presence of chelating agents and other external conditions which break ionic bonds. Thus, the most preferred crosslinking agents for use in the practice of the invention are those that form covalent bonds with the water-soluble polymer.
The crosslinking agent can be added to the aqueous polymer solution prior to emulsification, to the dispersion of water-soluble droplets in the oil phase, or, in some cases, to the inverted emulsion. The order of addition of the crosslinking agent will depend not only upon the particular polymer and crosslinking agent chosen but also upon the rate of the crosslinking reaction. An important consideration is that the rate of crosslinking not be such that it is competitive with the emulsification. Preferably, the crosslinking reaction is slow enough to permit complete emulsification of the aqueous and oil phases. Optionally, the crosslinking agent may be added to the aqueous polymer solution, and the pH of the solution adjusted to suitably slow down the rate of crosslinking prior to the solution being added to, and emulsified with, the oil phase. Subsequently, the pH is adjusted to provide the desired rate of crosslinking reaction.
Optionally, a catalyst can be added to initiate the crosslinking reaction. Choice of a particular catalyst will depend upon the particular water-soluble polymer selected, as well as the crosslinking agent, and other reaction conditions. The selection of any particular catalyst, in any case, can be readily done by those skilled in the art. Examples of catalysts that can be used in the practice of the invention include, but are not limited to, p-toluene sulfonic acid and hydrochloric acid. Optionally, heat may be provided as the catalyst in some cases.
The oil phase (i.e. the non-aqueous solvent phase) can be any inert hydrophobic liquid which can be readily separated from the dispersion of water-swellable polymer particles. In general, any hydrocarbon can be used as the oil phase liquid. Preferably, the hydrocarbon is toluene, o-xylene, or isooctane. Of concern, however, is that the crosslinking agent should not be soluble in the oil phase. Neither should the water-soluble polymer solution be soluble or miscible with the hydrocarbon used as the oil phase. Preferably, the hydrocarbon should also be substantially pure, although mixtures of hydrocarbons may be used. Preferably, the hydrocarbon is substantially non-volatile and non-polar. Those skilled in the art will be able to chose a suitable hydrocarbon for use in the practice of the invention. Although not critical to the invention disclosed herein, the hydrocarbon chosen, from a practical standpoint, should be low in cost.
In order to emulsify the water-soluble polymer phase into the oil phase to give a water-in-oil emulsion, one or more emulsifying agents of the water-in-oil type (a surfactant) are used in the amount of from about 0.010 to about 10.0 weight percent of the oil phase. Any commonly used water-in-oil emulsifying agent can be used in the practice of the invention, for example, hexadecyl sodium phthalate, sorbitan monostearate, metal soaps, and the like. Nevertheless, how well a given emulsifier works in any particular case depends upon the polymer solution to be emulsified, the composition of the oil phase, and the means of emulsification.
The emulsifier must function to stabilize the water-in-solvent system of the invention sufficiently to permit controlled crosslinking of the hydrophilic polymers to yield particles that are substantially homogeneous in size and shape, i.e. approximately 80% of the particles are within one standard deviation of the mean size, and at least approximately 95% of the particles are spherical. Otherwise, the aqueous phase particles will agglomerate and eventually form one continuous water-layer.
The choice of emulsifiers depends upon a number of factors including the water-soluble polymer selected and the size of the particles to be formed. It is important to match the properties of the emulsifier with the size and properties of the particles to be produced. Important emulsifier properties to be considered in this choice are the molecular length and charge of the emulsifier. Another important factor is the ability of the emulsifier to wrap different amounts of water molecules, allowing for the production of different size microspheres.
Still another important property in choosing an emulsifier is its ability to maintain a hydrophile-lipophile balance. This hydrophile-lipophile balance is the balance between the size and strength of the hydrophilic (water-loving or polar) and the lipophilic (oil-loving or non-polar) functional groups on the emulsifier.
Emulsifiers useful in the invention have low HLB (Hydrophile-Lipophile Balance) values, generally HLB values of less than approximately 8, preferably less than approximately 6, more preferably between about 4 and about 6. A particularly prefered emulsifier is SPAN 60. Those skilled in the art can readily determine, which water-in-oil emulsifier, i.e. surfactant., to use in any given case.
The crosslinking reaction is controlled to some extent by the particular reactants involved, the concentration of the reactants, the length of reaction time, the temperature, and the pH of the reaction mixture. The ratio of the weight of the water-soluble polymer to that of the crosslinking agent will depend upon the particular polymer and crosslinking agent employed. This ratio can vary from about 0.2 to about 200, preferably from about 1 to about 10.
The extent of crosslinking of the polymer droplets is generally controlled by controlling the length of the reaction time while maintaining the reaction at room temperature. In any particular case, the most suitable reaction time can be readily determined by those skilled in the art. In some cases the crosslinking reaction can be stopped by the addition of an alcohol, such as methanol or isopropanol, to the reaction mixture. The hydroxyl groups of the alcohol react with the functional groups of the crosslinking agent, quenching the reaction with the water-soluble polymer. The pH of the reaction mixture can be adjusted to control the rate of crosslinking, for example, by addition of a base, such as ammonium hydroxide, or an acid, such as acetic or hydrochloric acid, to the reaction mixture. The combination of reaction rate and reaction time determines the extent of crosslinking of the polymer droplets.
The extent of crosslinking is important to the present invention because it determines the rigidity and elasticity of the resulting water-swellable polymer particles. Elasticity is important because it allows the microspheres to be injected through a narrow guage needle. The elasticity of the microspheres is determined by the porosity and pore size. The porosity and pore size in turn are determined by the ratio of crosslinker to base polymer and by the homogeneity of the components within each microsphere. Optionally, a second surfactant can be employed to assist in obtaining and maintaining the homogeneity of the particles.
Unlike the emulsifier, this second surfactant is a solubilizer having an HLB between about 10 and about 18, preferably between about 12 and about 18, more preferably between about 14 and about 16. The use of such a surfactant is not essential, but promotes the homogeneity, elasticity, and clarity of the microspheres.
The extent and type of crosslinking is also important to the present invention because it prevents rapid breakdown of the microspheres in the body. Unlike, the process of Rothman, et al. (U.S. Pat. No. 4,124,705) the selection of water-soluble polymer and crosslinker allows for multiple types of linkages. Nonlimiting examples of the types of linkages formed by the process of the present invention are xcex1(1xe2x86x924), xcex1(1xe2x86x923), xcex1(1xe2x86x921), xcex2(1xe2x86x924), and xcex2(1xe2x86x926). In part, multiple types of linkages are formed because of the different functional groups on the water-soluble polymer. For example, different polysaccharides have different numbers of hydroxyl groups located at different positions on the sugar ring. Each of these hydroxyl groups has a different reactivity to the crosslinker and forms different types of linkages. Because the process of the invention can crosslink any of these hydroxyl groups, the process of the invention is capable of forming many different types of linkages within the same polymer. This is also true of water-soluble polymers having other types of functional groups, including polymers having multiple types of functional groups. The number and type of different linkages can be controlled through selection of the reaction components and parameters.
The extent of agitation of the emulsion during droplet formation depends not only upon the recipe of the reactants involved in any particular case but also upon the size of the particles to be formed. For example, as will become more apparent hereinafter, toluene containing SPAN 60 emulsifying agent, i.e. sorbitan monostearate having a HLB of about 4.7, which is available from ICI Americas, can be used to make microscopic-size particles, e.g., 50 micrometers in diameter, or submicroscopic-size particles, e.g., 0.2 micrometers in diameter, according to the concentration of the SPAN 60 and the degree of agitation.
In general terms, as the concentration of the emulsifier increases, the size of the particles formed decreases. This is because the emulsifier stabilizes the individual microspheres and inhibits their agglomeration. Similarly, as the degree of agitation increases, the size of the particles formed decreases. The prefered degree of agitation in the present invention is a moderate agitation of about 100 rpm to about 600 rpm by stirring, more preferably about 200 rpm to 400 rpm. This level of agitation is sufficient to form particles in the range of about 10 micrometers to about 250 micrometers and to mix the particles after formation without destroying them. If a lower level of agitation is used, larger particles will result. If the level of agitation becomes too low, the microspheres will agglomerate, forming large aggregates. On the other hand, if a higher level of agitation is used, submicron particles will result. It is not possible to control the uniformity of the size and/or shape of such submicron particles. These submicron particles are not advantageous for implants, soft tissue augmentation, or tissue scaffolding because they easily migrate to different organs or undergo endocytosis by inflammatory cells of the host, such as neutrophils and macrophages.
The separation of the crosslinked particles from the aqueous phase can be accomplished by various known procedures. Generally, after inversion of the polymer particles into an aqueous phase, the oil layer is separated from the aqueous phase, e.g., by sedimentation, leaving an aqueous dispersion of crosslinked polymer particles. Then, the crosslinked water-soluble polymer particles are separated from the aqueous phase by filtration, after which the particles are washed and dehydrated with methanol. The particles are then dried. This can be done by merely spreading the particles out on a flat surface; however, in the case of an industrial scale operation, the particles can be dried in a fluidized bed drier conventionally used for such a purpose.
The particles may be microcapsules, microspheres, or beads. Microcapsules are defined as polymer particles containing one or more encapsulated compositions. Preferably, the encapsulated compositions are drugs, for example, but not limited to, antigens, antibodies, growth factors, inhibitors, antibiotics, antisense oligonucleotides, antiviral compositions, anti-cancer compositions, therapeutic agents, and other compositions to be administered to a patient, or delivered to a specific site in a patient. Microcapsules are large enough to be seen by the naked eye. Microspheres, on the other hand, are much smaller particles that generally do not contain encapsulated materials. Moreover, they may require optical lo microscopy to be seen. Beads are spherically-shaped particles that are large enough to be seen with the naked eye. The limit of visibility of beads is in the range of from 60-100 micrometers in diameter.
The size of particles resulting from the practice of the invention, quite advantageously, can vary over a wide range of sizes, both microscopic and submicroscopic, according to the average droplet size of the emulsion, which depends upon the composition, type and concentration of emulsifier, as well as the emulsification procedure and conditions. The particles of the invention are generally between about 10 micrometers and about 250 micrometers in diameter. The polymer particles are preferably greater than about 10 micrometers and less than about 212 micrometers in diameter, more preferably greater than about 10 micrometers and less than about 150 micrometers in diameter. Advantageously, the particles formed are substantially homogeneous in size and shape. Another feature of the water-swellable polymer particles of the invention is that when dispersed in water and injected through a narrow guage hypodermic syringe needle (20-22 guage), a worm-like thread is formed. Injectability is an important chacteristic of the water-swellable microspheres when the particles are used for drug delivery or therapy.